Structure and sequence engineering approaches to improve in vivo expression of nucleic acid-delivered antibodies

Sep 30, 2026

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Monoclonal antibodies are widely used biologics, yet global access is constrained by challenges in production, storage, and distribution. The work investigates how antibody structure and sequence influence in vivo expression using a synthetic DNA platform with the SARS-CoV-2 antibody 2196 DNA-encoded monoclonal antibody (DMAb-2196). Through heavy/light chain “chain-swap” rational design and structural modeling, the study derives an antibody frequency score to predict expression-enhancing mutations. A single mutation can raise expression up to 2-fold, and mutation combinations further improve expression, enabling a generalized pipeline for therapeutic antibody delivery across indications.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages16

Reading Time40 min

AccessFree to download

Introduction

Rationale for antibody therapeutics and gene-encoded delivery

Platforms and expression timescales

Limitations tied to biologically relevant circulating levels

Original Article Structure and sequence engineering approaches to improve in vivo expression of nucleic acid-delivered antibodies Michaela Helble, 1,2 Jacqueline Chu, 1 Kaitlyn Flowers, 1 Abigail R. Trachtman, 1 Alana Huynh, 1 Amber Kim, 1 Nicholas Shupin, 1 Casey E. Hojecki, 1 Ebony N. Gary, 1 Shahlo Solieva, 1,3 Elizabeth M. Parzych, 1 David B. Weiner, 1,2 Daniel W. Kulp, 1,2,3 and Ami Patel 1 1Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA 19104, USA; 2Department of Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; 3Department of Biochemistry and Molecular Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA Monoclonal antibodies are an important class of biologics with over 160 Food and Drug Administration/European Unionapproved drugs. A signiﬁcant bottleneck to global accessibility of recombinant monoclonal antibodies stems from complexities related to their production, storage, and distribution. Recently, gene-encoded approaches such as mRNA, DNA, or viral delivery have gained popularity, but ensuring biologically relevant levels of antibody expression in the host remains a critical issue. Using a synthetic DNA platform, we investigated the role of antibody structure and sequence toward in vivo expression. SARS-CoV-2 antibody 2196 was recently engineered as a DNA-encoded monoclonal antibody (DMAb-2196). Utilizing an immunoglobulin heavy and light chain “chain-swap” methodology, we interrogated features of DMAb-2196 that can modulate in vivo expression through rational design and structural modeling. Comparing these results to natural variation of antibody sequences resulted in development of an antibody frequency score that aids in the prediction of expressionimproving mutations by leveraging antibody repertoire datasets. We demonstrate that a single amino acid mutation identiﬁed through this score increases in vivo expression up to 2-fold and that combinations of mutations can also enhance expression. This analysis has led to a generalized pipeline that can unlock the potential for in vivo delivery of therapeutic antibodies across many indications. INTRODUCTION Antibody therapeutics are currently a critical component of care across a variety of ﬁelds from cancer to autoimmunity to infectious disease. For cancer, checkpoint blockade antibodies nivolumab, pembrolizumab (anti-PD1), and ipilimumab (anti-CTLA-4) have achieved unprecedented clinical responses and sparked a revolution in cancer treatment 1–5; over 55 antibodies in the United States/ European Union have been approved for cancer treatment alone.6 In autoimmunity, anti-tumor necrosis factor (TNF) antibody adalimumab used in the treatment of ulcerative colitis and Crohn’s disease remains the top-selling monoclonal.7–9 The recent approval of nirsevimab for use in respiratory syncytial virus (RSV) shows the signiﬁcance of antibodies for infectious disease prevention.10, 11 This was also mirrored in the numerous anti-SARS-CoV-2 antibody development efforts initiated during the pandemic.12 These examples, and many others, showcase the importance of antibody therapies. They have revolutionized treatment across diverse ﬁelds and their continued development and regulatory approval remains critical. The recombinant monoclonal antibody (mAb) development process includes rigorous antibody biochemical and biophysical property analyses, quality control assessments, 13, 14 complexities related to dosage and route of administration, 15, 16 as well as other important manufacturing considerations. Gene-encoded delivery of antibodies in vivo may offer a way to circumvent these limitations associated with recombinant antibody production. There are several major platforms that make use of in vivo gene-encoded delivery, including viral vectors, mRNA-LNP, and DNA delivery. An antibody of interest is encoded and optimized for direct in

FAQs

What problem does the study address in nucleic acid-delivered antibodies?

It addresses the bottleneck of achieving biologically relevant in vivo antibody expression levels from nucleic acid delivery, which limits therapeutic effect.

How does the study interrogate antibody sequence effects on expression?

It uses a synthetic DNA platform and a heavy/light chain “chain-swap” methodology with rational design and structural modeling of DMAb-2196.

What output does the study provide to guide mutation selection?

An antibody frequency score that predicts expression-improving mutations using antibody repertoire datasets.

Sep 30, 2026

Upload by Jiven